pT3TS-zAsCpf1

107185 Danio rerio Bacterial Expression

Plasmid Information

Purpose

plasmid to carry out IVT of AsCpf1 (zebrafish codon optimized)

Depositor

Antonio Giraldez

Article

Moreno-Mateos et al Nat Commun. 2017 Dec 8;8(1):2024. doi: 1

Insert

AsCpf1

Tags

NLS-3xHA

Available Since

March 8, 2018

Availability

Available

Sequence Information

Length

7,217 bp

Topology

circular

Molecule Type

other DNA

View on AddGene.org

Features

3'UTR 1

Label	Gene	Product	Position	Strand
Xenopus globin 3'-UTR	—	—	4193..4319	Fwd
DNA Sequence AACCAGCCTCAAGAACACCCGAATGGAGTCTCTAAGCTACATAATACCAACTTACACTTTACAAAATGTTGTCCCCCAAAATGTAGCCATTCGTATCTGCTCCTAATAAAAAGAAAGTTTCTTCAC

5'UTR 1

Label	Gene	Product	Position	Strand
Xenopus globin 5'-UTR	—	—	43..86	Fwd
DNA Sequence CTTGTTCTTTTTGCAGAAGCTCAGAATAAACGCTCAACTTTGG

CDS 4

Label	Gene	Product	Position	Strand
AsCpf1	—	CRISPR-associated protein from Acidamin…	107..4031	Fwd
DNA Sequence ATGGGCACCCAGTTCGAGGGATTCACCAACCTGTACCAGGTGTCCAAGACCCTGAGATTCGAGCTGATCCCTCAGGGAAAGACCCTGAAGCACATCCAGGAACAGGGATTCATCGAAGAGGACAAGGCCAGAAACGACCACTACAAAGAGCTGAAGCCCATCATCGACCGCATCTACAAGACCTACGCCGACCAGTGCCTGCAGCTCGTGCAGCTGGACTGGGAGAACCTGAGCGCCGCCATCGACAGCTACAGAAAAGAGAAAACCGAGGAAACACGCAACGCCCTGATCGAAGAGCAGGCCACCTACAGAAACGCCATCCACGACTACTTCATCGGAAGAACCGACAACCTGACCGACGCCATCAACAAGAGACACGCCGAGATCTACAAGGGACTGTTCAAGGCCGAGCTGTTCAACGGAAAGGTGCTGAAGCAGCTGGGAACCGTGACCACCACCGAGCACGAGAACGCCCTGCTGAGAAGCTTCGACAAGTTCACCACCTACTTCAGCGGCTTCTACGAGAACAGAAAGAACGTGTTCAGCGCCGAGGACATCAGCACCGCCATCCCTCACAGAATCGTGCAGGACAACTTCCCCAAGTTCAAAGAGAACTGCCACATCTTCACCAGACTGATCACCGCCGTGCCTAGCCTGCGCGAGCACTTCGAGAACGTGAAGAAGGCCATCGGAATCTTCGTGTCTACCTCTATCGAGGAAGTGTTCAGCTTCCCATTCTACAACCAGCTGCTGACCCAGACCCAGATCGACCTGTATAATCAGCTGCTGGGAGGAATCAGCAGAGAGGCCGGAACCGAGAAGATCAAGGGACTGAACGAGGTGCTGAACCTGGCCATCCAGAAGAACGACGAGACAGCCCACATCATTGCCAGCCTGCCTCACAGATTCATCCCTCTGTTCAAGCAGATCCTGAGCGACAGAAACACCCTGAGCTTCATCCTGGAAGAGTTCAAGAGCGACGAGGAAGTGATCCAGAGCTTCTGCAAGTACAAGACACTGCTGAGAAACGAGAACGTGCTGGAGACAGCCGAGGCCCTGTTCAACGAGCTGAACAGCATCGACCTGACACACATCTTCATCAGCCACAAGAAGCTGGAGACAATCAGCAGCGCCCTGTGCGACCACTGGGACACCCTGAGAAACGCCCTGTACGAGAGAAGAATCAGCGAGCTGACCGGAAAGATCACCAAGAGCGCCAAAGAGAAGGTCCAGAGAAGCCTGAAGCACGAGGATATCAACCTGCAGGAAATCATCAGCGCCGCTGGAAAAGAGCTGAGCGAGGCCTTCAAGCAGAAAACCAGCGAGATCCTGTCTCACGCCCACGCCGCCCTGGACCAGCCTCTGCCTACCACCCTGAAGAAGCAGGAAGAGAAAGAGATCCTGAAAAGCCAGCTGGACAGCCTGCTGGGACTGTACCATCTGCTGGATTGGTTCGCCGTGGACGAGAGCAACGAGGTGGACCCTGAGTTCAGCGCCAGACTGACCGGAATCAAGCTGGAGATGGAGCCTAGCCTGAGCTTCTACAACAAGGCTAGAAACTACGCCACCAAGAAACCCTACAGCGTGGAGAAGTTCAAACTGAACTTCCAGATGCCTACCCTGGCCAGCGGATGGGACGTGAACAAAGAGAAGAACAACGGCGCCATCCTGTTCGTGAAGAACGGACTGTACTACCTGGGAATCATGCCTAAGCAGAAGGGAAGATACAAGGCTCTGAGCTTCGAGCCTACCGAGAAAACAAGCGAGGGATTCGACAAGATGTACTACGATTACTTCCCTGACGCCGCCAAGATGATCCCTAAGTGCAGCACCCAGCTGAAGGCCGTGACCGCCCACTTCCAGACCCACACCACCCCTATCCTGCTGAGCAACAACTTCATCGAGCCTCTGGAGATCACCAAAGAGATCTACGACCTGAACAACCCCGAGAAAGAGCCTAAGAAGTTCCAGACCGCCTACGCCAAGAAAACCGGCGACCAGAAGGGATACCGCGAGGCTCTGTGCAAGTGGATCGACTTCACCAGAGACTTTCTGAGCAAGTACACCAAGACCACCAGCATCGATCTGAGCAGCCTGCGCCCTAGCAGCCAGTACAAGGACCTGGGAGAGTACTACGCCGAGCTGAACCCTCTGCTGTACCACATCAGCTTCCAGAGAATCGCCGAGAAAGAGATTATGGACGCCGTGGAGACAGGCAAGCTGTACCTGTTCCAGATCTACAACAAAGACTTCGCCAAGGGACACCACGGAAAGCCTAACCTGCACACCCTGTACTGGACCGGACTGTTCAGCCCTGAGAACCTGGCTAAGACCAGCATCAAGCTGAACGGACAGGCTGAGCTGTTCTACAGACCTAAGAGCAGAATGAAGAGAATGGCCCACAGACTGGGAGAGAAGATGCTGAACAAGAAACTGAAGGACCAGAAAACCCCTATCCCCGACACACTGTACCAGGAACTGTACGACTACGTCAACCACAGACTGAGCCACGACCTGTCTGACGAGGCCAGAGCCCTGCTGCCTAACGTGATCACAAAAGAGGTGTCCCACGAGATCATCAAGGACAGACGCTTCACCAGCGACAAGTTCTTTTTCCACGTGCCCATCACCCTGAACTACCAGGCCGCCAACAGCCCTAGCAAGTTCAACCAGAGAGTGAACGCCTACCTGAAAGAGCACCCTGAGACACCTATCATCGGAATCGACAGAGGCGAGCGCAACCTGATCTACATCACCGTGATCGACAGCACCGGAAAAATCCTGGAGCAGAGATCCCTGAACACCATCCAGCAGTTCGACTACCAGAAGAAACTGGACAATCGCGAGAAAGAGCGCGTCGCCGCCAGACAGGCTTGGAGCGTCGTGGGAACCATCAAGGACCTGAAGCAGGGATACCTGAGCCAGGTCATCCACGAGATCGTGGACCTGATGATCCACTACCAGGCTGTGGTGGTGCTGGAGAATCTGAACTTCGGATTCAAGAGCAAGCGCACCGGAATCGCTGAGAAGGCCGTGTACCAGCAGTTTGAGAAAATGCTGATCGACAAGCTGAACTGCCTGGTGCTGAAGGACTACCCTGCCGAGAAAGTGGGAGGCGTGCTGAACCCTTACCAGCTGACCGACCAGTTCACCAGCTTCGCCAAGATGGGAACCCAGAGCGGATTCCTGTTCTACGTGCCAGCCCCTTACACCAGCAAGATCGACCCTCTGACCGGATTCGTGGACCCTTTCGTGTGGAAAACCATCAAGAACCACGAGAGCAGAAAGCACTTCCTGGAGGGCTTCGACTTCCTGCACTACGACGTGAAAACAGGCGACTTCATCCTGCACTTCAAGATGAACAGAAACCTGAGCTTCCAGCGCGGACTGCCTGGATTCATGCCTGCCTGGGACATCGTGTTCGAGAAAAACGAGACACAGTTCGACGCCAAGGGAACCCCTTTTATCGCCGGAAAAAGAATCGTGCCTGTGATCGAGAACCACAGATTCACCGGAAGATACAGAGATCTGTACCCCGCTAACGAGCTGATCGCCCTGCTGGAAGAGAAGGGAATCGTGTTCCGCGACGGAAGCAACATCCTGCCTAAGCTGCTGGAGAACGACGACAGCCACGCTATCGACACAATGGTGGCCCTGATCAGATCTGTGCTGCAGATGAGAAACAGCAACGCCGCCACCGGCGAGGACTACATCAACAGCCCTGTGCGCGACCTGAACGGCGTGTGCTTCGACAGCAGATTCCAGAACCCTGAGTGGCCTATGGACGCTGACGCCAACGGCGCCTACCATATCGCCCTGAAGGGACAGCTGCTGCTGAACCATCTGAAAGAGAGCAAAGACCTGAAGCTGCAGAACGGAATCAGCAACCAGGACTGGCTGGCCTATATCCAGGAGCTGAGAAAC Protein Sequence MGTQFEGFTNLYQVSKTLRFELIPQGKTLKHIQEQGFIEEDKARNDHYKELKPIIDRIYKTYADQCLQLVQLDWENLSAAIDSYRKEKTEETRNALIEEQATYRNAIHDYFIGRTDNLTDAINKRHAEIYKGLFKAELFNGKVLKQLGTVTTTEHENALLRSFDKFTTYFSGFYENRKNVFSAEDISTAIPHRIVQDNFPKFKENCHIFTRLITAVPSLREHFENVKKAIGIFVSTSIEEVFSFPFYNQLLTQTQIDLYNQLLGGISREAGTEKIKGLNEVLNLAIQKNDETAHIIASLPHRFIPLFKQILSDRNTLSFILEEFKSDEEVIQSFCKYKTLLRNENVLETAEALFNELNSIDLTHIFISHKKLETISSALCDHWDTLRNALYERRISELTGKITKSAKEKVQRSLKHEDINLQEIISAAGKELSEAFKQKTSEILSHAHAALDQPLPTTLKKQEEKEILKSQLDSLLGLYHLLDWFAVDESNEVDPEFSARLTGIKLEMEPSLSFYNKARNYATKKPYSVEKFKLNFQMPTLASGWDVNKEKNNGAILFVKNGLYYLGIMPKQKGRYKALSFEPTEKTSEGFDKMYYDYFPDAAKMIPKCSTQLKAVTAHFQTHTTPILLSNNFIEPLEITKEIYDLNNPEKEPKKFQTAYAKKTGDQKGYREALCKWIDFTRDFLSKYTKTTSIDLSSLRPSSQYKDLGEYYAELNPLLYHISFQRIAEKEIMDAVETGKLYLFQIYNKDFAKGHHGKPNLHTLYWTGLFSPENLAKTSIKLNGQAELFYRPKSRMKRMAHRLGEKMLNKKLKDQKTPIPDTLYQELYDYVNHRLSHDLSDEARALLPNVITKEVSHEIIKDRRFTSDKFFFHVPITLNYQAANSPSKFNQRVNAYLKEHPETPIIGIDRGERNLIYITVIDSTGKILEQRSLNTIQQFDYQKKLDNREKERVAARQAWSVVGTIKDLKQGYLSQVIHEIVDLMIHYQAVVVLENLNFGFKSKRTGIAEKAVYQQFEKMLIDKLNCLVLKDYPAEKVGGVLNPYQLTDQFTSFAKMGTQSGFLFYVPAPYTSKIDPLTGFVDPFVWKTIKNHESRKHFLEGFDFLHYDVKTGDFILHFKMNRNLSFQRGLPGFMPAWDIVFEKNETQFDAKGTPFIAGKRIVPVIENHRFTGRYRDLYPANELIALLEEKGIVFRDGSNILPKLLENDDSHAIDTMVALIRSVLQMRNSNAATGEDYINSPVRDLNGVCFDSRFQNPEWPMDADANGAYHIALKGQLLLNHLKESKDLKLQNGISNQDWLAYIQELRN
nucleoplasmin NLS	—	bipartite nuclear localization signal f…	4031..4079	Fwd
DNA Sequence AAAAGACCTGCCGCTACCAAGAAGGCCGGACAGGCCAAGAAGAAGAAG Protein Sequence KRPAATKKAGQAKKKK
3xHA	—	three tandem HA epitope tags	4085..4166	Fwd
DNA Sequence TACCCTTACGACGTGCCAGACTACGCCTACCCATACGATGTGCCTGATTATGCTTACCCATATGACGTGCCCGATTACGCC Protein Sequence YPYDVPDYAYPYDVPDYAYPYDVPDYA
AmpR	bla	beta-lactamase	5175..6036	Fwd
DNA Sequence ATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAA Protein Sequence MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW

promoter 4

Label	Gene	Product	Position	Strand
T3 promoter	—	—	0..19	Fwd
DNA Sequence AATTAACCCTCACTAAAGG
T7 promoter	—	—	4405..4424	Rev
DNA Sequence TAATACGACTCACTATAGG
AmpR promoter	bla	—	5070..5175	Fwd
DNA Sequence CGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCTGCTCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGT
lac promoter	—	—	7118..7149	Fwd
DNA Sequence TTTACACTTTATGCTTCCGGCTCGTATGTTG

protein_bind 2

Label	Gene	Product	Position	Strand
CAP binding site	—	—	7082..7104	Fwd
DNA Sequence TAATGTGAGTTAGCTCACTCAT
lac operator	—	—	7156..7173	Fwd
DNA Sequence TTGTGAGCGGATAACAA

rep_origin 2

Label	Gene	Product	Position	Strand
f1 ori	—	—	4588..5044	Rev
DNA Sequence ACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTT
ori	—	—	6206..6795	Fwd
DNA Sequence TTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAA